Although the rate of protein breakdown may determine the size of tissues as well as the levels of specific proteins, previous studies of the control of this process in cultured cells have suffered from various limitations. Because serum contains an undefined mixture of mitogens, hormones, and binding proteins, the effects of these factors on protiolysis have often been determined in cells deprived of serum, a highly unphysiologic, toxic, and ultimately lethal condition that accelerates proteolysis and produces negative protein balance. Other studies have manipulated growth rates by varying cell population density. However, density-inhibited and subconfluent cultures differ not only in growth rate, but in the extent of cell-cell contact and the availability of certain nutrients. We will use more selective methods to manipulate the proliferative rate of cells to investigate the relations between proteolysis and growth rate, nutrient supply, and certain physiologically important hormones in vascular smooth muscle and Balb/c-eT3 cells. We will use a defined hormone-supplemented medium that will maintain viability and neutral nitrogen balance of cells without promoting cell division, that would permit precise control of hormone and nutrient levels. We will then add mitogens to this defined "nongrowth" medium to manipulate proliferative rate selectively, independent of cell population density, in single variable experiments in which the control, non-growing cells are not in an unphysiologic catabolic state. We will then test these hypotheses: (1) Altered rates of protein breakdown contribute to the accumulation of protein in cells proliferating in response to serum-associated mitogens. (2) The population density of cell cultures does not regulate protein breakdown or synthesis directly, but by effects on nutrient supply or growth rate. (3) Certain hormones found in serum may contribute to the physiologic regulation or protein balance nd perhaps to the control of cell growth. We will complement the studies of overall protein breakdown with parallel measurements of fucoglycoprotein catabolism. Also, using cultures of contractile vascular smooth muscle cells we will determine if increased rates of catabolism account for the loss of actin that seems to accompany the modulation of these cells to the "synthetic" phenotype, that may be an early event in atherogenesis.